Some forms of Alzheimer's disease are directly related to the enhanced production by gamma-secretase of a long form of the amyloid beta peptide relative to less toxic shorter forms (long form = AB42; short form = AB40). It is also known that certain small drug-like molecules can also perturb the normal AB42:AB40 production ratio. In this project we will provide a first-ever structural characterization of the protein precursor from which both long and short forms of the amyloid-B peptide are derived via y-secretase cleavage: the N- terminal 672-770 fragment of the amyloid precursor protein (APP672-770). Both wild type APP672-770 and known disease-linked mutant forms of this protein will be characterized. By analyzing, comparing, and contrasting the structures of the different forms of APP672-770 we will be able to formulate experimentally-testable hypotheses for exactly how and why amyloid-beta peptide production goes awry. We will also characterize the complex formed between APP672-770 and a small molecule known to modulate its cleavage by gamma-secretase. Insight from these studies may ultimately be used as the basis for developing novel therapeutic agents that are designed either to prohibit or to correct the aberrant processing of the amyloid precursor that occurs in some forms of Alzheimer's disease. In preliminary data we have succeeded in expressing and purifying both wild type and disease-linked mutant forms of APP672-770 and have shown that high quality NMR spectra can be obtained. The specific aims are: (1) Characterize the 3-D structure, dynamics, and membrane interactions of wild type APP672-770 using NMR spectroscopy and other methods. APP672-770 is a transmembrane protein that serves as the substrate for gamma- secretase cleavage to produce the amyloid-beta peptide. (2) Characterize the 3-D structures, dynamics, and membrane interactions of mutant forms of APP672-770 that have been linked to familial Alzheimer's disease (FAD). Results will be compared to those for the wild type protein in order to illuminate the structural biophysical basis for how mutations promote Alzheimer's disease. (3) Characterize the binding of a small molecule to APP672-770 that both inhibits gamma-secretase cleavage and lowers the AB42:AB40 production ratio. The molecule of interest has previously been shown to act through pre-association with APP672-770 rather than through association with gamma-secretase. [unreadable] [unreadable] [unreadable]